Streptolysin O (SO) is a hemolytic protein produced by most group A streptococci. One property of SO is its antigenicity with the result that antistreptolysin O (ASO) is usually produced in response to Streptococcus A infections. While ASO binds SO and inhibits SO hemolytic activity, the amount of ASO produced in response to the infection may be in excess of the amount required to inhibit the SO. Moreover, repeat infections frequently trigger an amplified immune response and excess production of ASO. Because ASO also bonds to M type proteins, this leads to secondary immune responses, the sequellae of which include degeneration or destruction of certain heat and kidney tissues which may result in rheumatic fever or acute glomorulonephritis.
It is also well known that SO hemolytic activity is neutralized or destroyed in its oxidized state. Since the SO antigen is readily available commercially and because of the described properties of SO and ASO, the measurement and monitoring of a patient ASO titer has been a feasible and useful diagnostic tool.
Most commercially available kits for determining ASO titer are based on the Rantz and Randall methodology Proc. Sec. Exp. Biol. Med. (N.Y.) 59,22 (1945). The procedure calls for the technician to draw several mls of blood, and then process it into serum. The serum is then diluted and incubated at 37.degree. C. with SO. After an initial incubation period of 15 minutes, a constant amount of 5% washed rabbit, sheep, or human type O red blood cells (RBC's) are added, and the samples are again incubated. ASO titer is determined about 45 minutes later by noting which tubes contain lysed RBC's. The procedure is very time consuming and tedious. It also has several steps at which pipetting errors can easily occur and cause inaccurate results. In addition, it requires that the technician have access to a centrifuge, an incubator, and a fresh supply of washed 5% RBC's.
A more recent procedure is described by Ricci et al., j'l Clin. Microb., Vol. 8, no. 3, 263-267 (1978) and in U.S. Pat. No. 4,148,609 issued Apr. 10, 1979. In the Ricci method SO is chemically altered through oxidation to form disulfide bonds, chemically inactivating the hemolytic activities of the molecules. After incubating the oxidized SO with a blood sample possibly containing ASO, the SO is once again chemically altered with a reducing substance to reactivate the SO hemolytic activity. The SO in excess of that necessary to bind all the available ASO is then free to lyse the erythrocytes in the blood sample.
In most of these tests for determining ASO, the measurements are expressed in Todd Units. Thus one Todd Unit (TU) of SO has been arbitrarily defined as the amount of SO needed to completely lyse 1 ml of 5% red blood cells in saline in one hour at 37.degree. C. Correlatively one TU of ASO has been defined as that amount of ASO which bonds to 21/2 TU of SO. Whether one uses Todd Units or any other unit, arbitrary or otherwise, correlating the hemolytic activity of SO with its ability to bind with ASO, however, is a matter of choice, so long as such expressions or measurements are related to the observed ASO values of normal and diseased patients.
In this disclosure the term "hemolytic activity" has been used and is defined as the ability of SO to lyse blood cells; and the term "hemolytic capacity" has been used and is defined in relation to the amount of hemolytic activity and correlatively the amount of binding ability to ASO possessed by SO.